The transcriptional activity of the vitamin D receptor (VDR) is regulated by a number of coactivator and corepressor complexes, which bind to the VDR in a ligand (1,25(OH)2D3) dependent (coactivators) or inhibited (corepressors) process. In the keratinocyte the major coactivator complexes include the vitamin D interacting protein (DRIP) complex and the steroid receptor coactivator (SRC) complexes. Of the three SRC proteins critical for formation of SRC complexes, SRC3 plays the dominant role in keratinocytes. These coactivator complexes are not interchangeable in their regulation of keratinocyte proliferation and differentiation. We found that the DRIP complex is the main complex binding to VDR in the proliferating keratinocyte, whereas SRC3 and its associated proteins is the major coactivator binding to VDR in the differentiated keratinocyte. Moreover, we have found a specific role for DRIP205 in the regulation of wnt signaling pathways regulating keratinocyte proliferation, whereas SRC3 uniquely regulates the ability of 1,25(OH)2D3 to induce more differentiated functions such as lipid synthesis and processing required for permeability barrier formation and the innate immune response triggered by disruption of the barrier. These findings provide a basis by which we can understand how one receptor (VDR) and one ligand (1,25(OH)2D3) can regulate a large number of genes in a sequential and differentiation specific fashion. Furthermore, these observations indicate the potential for discovering small molecules that by selectively modulating the interactions between the coactivators and VDR can manifest specificity in regulating VDR function not achievable by ligands such as 1,25(OH)2D3 and its analogs. Hypothesis: The two main coactivator complexes, DRIP and SRC, differentially modulate the ability of VDR, its ligand 1,25(OH)2D3 and their interactions with 2-catenin to inhibit keratinocyte proliferation, while promoting keratinocyte differentiation leading to a protective epidermal barrier and normal hair follicle cycling. To test this hypothesis we have developed mice null for either DRIP205 or SRC3 that enable us to achieve the following aims. Aim 1 Determine the selective roles of DRIP205 and SRC3 in VDR regulation of epidermal and hair follicle proliferation, differentiation, and protective barrier formation. Aim 2. Determine the means by which DRIP205 and SRC3 modulate VDR/2-catenin interactions in their regulation of keratinocyte proliferation and differentiation of the epidermis and hair follicle. Aim 3. Develop the means to selectively inhibit DRIP205 and SRC3 modulation of VDR function. Significance: Understanding the mechanisms by which 1,25(OH)2D3 regulates important functions of the epidermis including its proliferation and development of a protective barrier and developing the means to selectively modulate these functions is expected to have important therapeutic applications including the prevention of skin cancer and the development of treatment for a variety of skin diseases manifesting as hyperproliferation and disrupted differentiation. PUBLIC HEALTH RELEVANCE: The mechanisms by which 1,25(OH)2D3 and its receptor VDR regulate keratinocyte differentiation and function remain unclear. We have discovered that different coactivators, DRIP and SRC in particular, regulate the transcriptional activity of VDR differently for different genes, and thus selectively affect different functions in the keratinocyte. By developing mice lacking either DRIP205 or SRC3, we will further explore these differences in an effort to more fully understand VDR transcriptional activity in the skin and develop means of selectively regulating these processes therapeutically.